176 EXPRESSION OF INSULIN-LIKE GROWTH FACTOR FAMILY MEMBERS AND GROWTH DIFFERENTIATION FACTOR 9 ON NELORE AND ANGUS HEIFERS WITH A LOW AND HIGH OVARIAN FOLLICLE COUNT

Abstract

Members of the IGF family are key intra-ovarian regulators of follicle growth, selection, and atresia. Growth differentiation factor 9 (GDF9) induces follicular somatic cells to undergo mitosis and differentiation during follicular development. Cattle from Bos indicus breeds are slower to reach sexual maturity and have longer calving intervals when compared with Bos taurus breeds (Luna-Nevarez et al. 2011). On the other hand, indicine cattle have greater number of ovarian follicles recruited per oestrous cycle when compared with taurine breeds (Alvarez et al. 2000). Our objective was to evaluate the expression of IGF1, IGFR1, IGF2, and GDF9 genes in follicles dissected from Nelore and Angus heifers with high (HFC) and low (LFC) follicle counts. Eighteen Nelore heifers and 22 Angus heifers (≈24 months old) were kept on Brachiaria brizantha grass and fed with a mix of grains with minerals and water ad libitum. Oestrous cycle was synchronized with 2 doses of PGF2α 11 days apart. Heifers were scanned with an ultrasound device (US; Mindray Vet DPS 2200, São Paulo, Brazil) with a 7.5-MHz probe 1 day after ovulation for 3 consecutive oestrous cycles. Animals were slaughtered ≈24 h after ovulation of the third cycle; 3 follicles from 2 to 4 mm in diameter were dissected from the ovary contralateral to the CL. Total RNA was extracted using the RNeasy Microarray Tissue Mini Kit (Qiagen, Valencia, CA, USA). Gene expression was evaluated using oligo-dT reverse transcription, Sybr Green Master Mix and Step One Plus (AB, Foster City, CA, USA) Real-Time PCR Detection System. Samples were analysed in duplicates and CT values were normalized to the housekeeping gene PPIA using the ΔCT method. Results were analysed using the PROC MIXED procedure of SAS with follicle as the repeated measure and cow as the subject. Individual differences were analysed using contrast (SAS 9.2). The effects of group (LFC × HFC), breed, and follicle diameter on mNRA abundance were tested. Follicle LSmean was higher (P < 0.05) in Nelore heifers (32 ± 3.1; LFC = 18; HFC = 52) when compared with Angus heifers (20 ± 2.6; LFC = 10; HFC = 27). Follicle diameter did not differ between breeds or groups. Expression of IGF1 and GDF9 was not different between follicles from Nelore and Angus heifers or between groups. Expression of IGFR1 was higher in follicles from Angus heifers (fold change 1.74; P < 0.04) when compared with follicles derived from Nelore heifers but not different between HFC and LFC groups within each breed. Expression of IGF2 mRNA was also higher (fold change 1.70; P < 0.04) in Angus heifers but not different between HFC and LFC groups within each breed. In conclusion, the higher expression of IGFR1 and IGF2 in Angus early antral follicles might be involved in important reproductive traits commonly observed in Bos taurus breeds. However, expression of IGF1, IGFR1, IGF2, and GDF9 in the follicle are not involved in the mechanisms determining different number of follicle recruited through the oestrous cycle in the same breed. This research and scholarship for Loureiro, Ereno, Favoureto, and Pupulim was from FAPESP.